LIGAND PHARMACOLOGY OF ESTROGEN RECEPTORS

该项目的长期目标是在分子水平上了解调节雌激素受体(ERα和ERβ)的配体的组织选择性。针对雌激素信号通路的药物，如他莫昔芬和雷洛昔芬，在某些组织中显示出抗雌激素效应，在其他组织中显示出雌激素效应。了解这种组织选择性的分子基础将有助于设计更安全和更有效的基于治疗的方法。所提出的实验旨在检验这一假设，即ER配体药理来自于三个参数的相互作用：配体结构、受体亚型(ERα和ERβ)以及特定基因启动子中作为ER调节部位的反应元件的类型。ER/AP-1位点是一个反应元件，观察到ERα和ERbeta的不同配体激活谱。具体目的是定位介导这种差异配体激活的ERpha和ERbeta区域。该项目的第二个具体目标是探索雷洛昔芬和他莫昔芬之间的结构差异，以确定这些药物的化学亚结构与观察到的R/AP-1位点配体激活的差异相关。ER/AP-1实验是一种人工细胞转录实验，它使用测试启动子驱动报告基因的转录，有助于发现ERa和ERbeta的差异配体激活特性。现在人们很感兴趣的问题是，靶组织中的内源性基因是否可以以类似的方式进行调节；即在表达ERbeta的组织中，哪些基因被抗雌激素如他莫昔芬和雷洛昔芬上调，哪些基因被雌二醇下调？通过使用差异基因表达阵列技术的实验，这个问题在特定目标3中得到了解决。用这种方法发现的内源性雌激素调节基因将通过特定目的4中描述的实验来表征。特别是，这些基因的启动子将被分析，以确定通过ER促进调节的反应元件的类型。

The long-term objective of this project is to develop a molecular-level understanding of the tissue selectivity of ligands that modulate the estrogen receptors (ERalpha and ERbeta). Drugs that target estrogen signaling pathways such as tamoxifen and raloxifene display antiestrogenic effects in certain tissues ans estrogenic effects in other tissues. Understanding the molecular basis of this tissue selectivity will be useful for designing safer and more effective-based therapeutics. The proposed experiments are designed to test the hypothesis that ER ligand pharmacology derives from the interplay of three parameters: ligand structure, receptor subtype (ERalpha and ERbeta), and the type of response element in the promoter of a given gene that is the site of regultion by the ER. The ER/AP-1 site is a response element where differential ligand activation profiles are observed for ERalpha and ERbeta. The specific aim is to map the regions of ERalpha and ERbeta that mediate this differential ligand activation. The second specific aim of the project is to probe the structural differences between raloxifene and tamoxifen to identify the chemical substructures of these drugs that correlate with the differences observed in ligand activation at an R/Ap-1 site. The ER/AP-1 assay that facilitated discovery of the differential ligand activation properties of ERalpha and ERbeta is an artificial cellular transcription assay that uses a test promoter driving transcription of a reporter gene. There is great interest now to ask whether endogenous in target tissues can be regulated in a similar way; i.e. in an ERbeta expressing tissue which genes are up-regulated by antiestrogens such as tamoxifen and raloxifene and down-regulated by estradiol? This question is addressed in specific aim 3 through experiments using differential gene expression array technology. The endogenous estrogen-regulated genes that are discovered with this approach will be characterized through experiments described in specific aim 4. In particular, the promoters of these genes will be analyzed to identify the type of response element that facilitates regulation through the ER.